This proposal is motivated by the need to assess microscopic viscoelastic properties, which have emerged as a powerful biomarker for a number of diseases, such as cancer, atherosclerosis, sickle cell disease, etc., but also have been identified as a driving force for many biological processes, such as carcinogenesis, angiogenesis, morphogenesis, etc. The emergence of novel biomaterials for regenerative medicine also calls for a better understanding of biomechanical cellular-level interactions. In the past, assessment of elastic properties of tissues was mostly limited to large-scale imaging using ultrasound and magnetic resonant imaging and to nanoscopic contact assessment using either optical tweezers or atomic force microscopy instruments, which paved the way to our better understanding of viscoelastic properties of cells and tissues and their importance for biomedical research. In the same time, it is now realized that there is a substantial technology gap in instrumentation capable of assessing non-invasively viscoelastic properties on a microscopic scale with high enough spatial resolution, high sensitivity and high speed. Recently, optical coherence elastography was successfully developed to assess elastic properties of tissues on the scale of 15-100 ??. Ideally, such an instrument should be fully compatible with existing instrumentation using fluorescence and Raman microscopy systems to provide an additional capability to those. Brillouin microscopy is emerging as a powerful tool for non-invasive biomedical imaging. Developing it into a powerful instrument for biomedical research and, potentially, clinical applications is considered to be the overarching goal of this proposal. Two strategies will be pursued through this grant application. The first approach is relying on spontaneous Brillouin microscopy, which is simpler in use, and, with relatively minor modifications, can be implemented as an option in already existing commercial fluorescent or Raman microscopes for a large biomedical community. The second strategy is to utilize nonlinear Brillouin spectroscopy and microscopy to boost the efficiency of the signal and data acquisition rate by astonishing 5 orders of magnitude. This methodology utilizes ultrashort pulse excitation and is fully compatible with multiphoton fluorescence microscopy, second- and third-harmonic microscopies and coherent anti-Stokes Raman microscopy. An additional benefit of nonlinear Brillouin microscopy is improved sectioning capabilities. The overall strategy is to design, construct and characterize both microscopes in parallel, since each of those offers distinct advantages for a particular set of applications, and to demonstrate their imaging capabilities for biologically relevant systems to image cells growth and development in response to a local viscoelastic environment and image developing zebrafish embryo during the first 72 hours post fertilization.